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Dynamic interaction of Y RNAs with chromatin and initiation proteins during human DNA replication

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journal contribution
posted on 17.12.2020, 06:27 by AT Zhang, AR Langley, CP Christov, E Kheir, Thomas ShafeeThomas Shafee, TJ Gardiner, T Krude
Non-coding Y RNAs are required for the initiation of chromosomal DNA replication in mammalian cells. It is unknown how they perform this function or if they associate with a nuclear structure during DNA replication. Here, we investigate the association of Y RNAs with chromatin and their interaction with replication proteins during DNA replication in a human cell-free system. Our results show that fluorescently labelled Y RNAs associate with unreplicated euchromatin in late G1 phase cell nuclei before the initiation of DNA replication. Following initiation, Y RNAs are displaced locally from nascent and replicated DNA present in replication foci. In intact human cells, a substantial fraction of endogenous Y RNAs are associated with G1 phase nuclei, but not with G2 phase nuclei. Y RNAs interact and colocalise with the origin recognition complex (ORC), the pre-replication complex (pre-RC) protein Cdt1, and other proteins implicated in the initiation of DNA replication. These data support a molecular 'catch and release' mechanism for Y RNA function during the initiation of chromosomal DNA replication, which is consistent with Y RNAs acting as replication licensing factors. © 2011. Published by The Company of Biologists Ltd.

Funding

We thank Cath Green and Sebastian Klinge for critical reading of this manuscript, Rachael Walker for help with flow cytometry and Stephen P. Jackson, Aloys Schepers, Michael Leffak, Stephen D. Bell, Heinz-Peter Nasheuer, Frank Grosse and Anindya Dutta for reagents. This work was supported by Cancer Research UK (project grant C1471/A8448 and vacation student bursary C1471/A10308) and by the Association for International Cancer Research (AICR Project Grant No: 10-0570). A. R. L. and T.J.G. were supported by Research Studentships from the Medical Research Council (MRC). The University of Cambridge Department of Zoology confocal suite was financed by the Wellcome Trust and the Newton Trust. Deposited in PMC for release after 6 months.

Cancer Research UK | C1471/A8448

Cancer Research UK | C1471/A10308

Association for International Cancer Research (AICR) | 10-0570

Medical Research Council (MRC)

Wellcome Trust

Newton Trust

Worldwide Cancer Research | 10-0570

History

Publication Date

01/01/2011

Journal

Journal of Cell Science

Volume

124

Issue

12

Pagination

(p. 2058-2069)

Publisher

The Company of Biologists

ISSN

0021-9533

Rights Statement

The Author reserves all moral rights over the deposited text and must be credited if any re-use occurs. Documents deposited in OPAL are the Open Access versions of outputs published elsewhere. Changes resulting from the publishing process may therefore not be reflected in this document. The final published version may be obtained via the publisher’s DOI. Please note that additional copyright and access restrictions may apply to the published version.